2. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
• Cova dos Santos or Cova da Meiga, is a karstic cave located
in Abadín (Lugo, NW of Spain) on the NW slope of the
Serra de Lourenzá, at an altitude of 360 m above see level and
25 m above the river Valiñadares, a tributary of the river,
which flows into the Bay of Biscay approximately 20 km to
the north.
• It is an area that may have been the natural pathway between
the faunas of the Cantabrian coast and those located further
south.
• This area is characterised by narrow
bands of limestone (Cándana and
Vegadeo limestone formations, Lower
Cambrian age) in which there are
caves of varying magnitude, but which
had so far not been prospected for the
location of palaeontological sites
3. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
• The cave has an horizontal extension of 198 m
and a total vertical development of 18.6 m.
• The topographical layout shows a “network
maze” type cavity pattern.
• The main room of the cavity is placed on the
central N-S axis. It has a wide vault, with a
relatively hemispherical section. The floor of this
room is made up of large blocks and fallen slabs,
on which fine sediments have accumulated.
4. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
The bone remains have been found in 4 areas:
• Area-01: the first bone remains recovered in
the cave (some cave bear remains) appeared in
a window connected with the central room.
• Area-02: in the main room, some fragmented
bones of small sized herbivores appeared
between the blocks.
• Area-03: the most important area of the cavity
in terms of number of remains. Bones of
medium and large-sized vertebrates, notably a
leopard tooth, several ursid metapodials and
remains of a large-sized mammal have been
found.
• Area-04: some herbivores remains.
5. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
• Three panels with bioglyphs (bear claw marks)
have been found in the cave.
6. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
• The skeletal remains studied (a total of 50 bones or bone fragment) come from the
collection of surface materials carried out during the topographical works in the cave.
46%
54%
Morphological identification
Unidentified
Cova dos Santos bone assemblage
Bos taurus
Felis catus
Ovis aries
Panthera pardus
Ursus arctos
Ursus spelaeus
What to do with these
unidentified remains?
7. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
• How to study a bone collection that is small and fragmented? The best way is to
apply molecular palaeontology techniques:
Bone
assemblage
Morphological
identification
Metric study
Imposible to
identify
ZooMS
Stable isotopes
Radiocarbon
dating
In this way we can select bones for:
8. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
Morphological
and
metric analysis
9. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
Bos taurus
• The remains morphologically attributable to Bos
taurus are a fragment of maxilla (SAN-04), the
distal fragment of a humerus (SAN-43), a
petrous bone (SAN-7) a vertebra (SAN-44) and
a calcaneus (SAN-50).
• The teeth and the humerus resembles those of medieval domestic cattle
from A Tara, and are even smaller. A Tara cows were described as
domestic animals of early medieval times and very small in size, as is still
common today in primitive breeds adapted to mountain environments (like
cachena breed in the picture).
10. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
Ovicaprines • The remains of ovicaprines are 1 right metacarpal (SAN-10), 1 juvenile
left tibia (SAN-11), 2 paired juvenile radios (SAN-12, left, and SAN-13,
right), 1 femur proximal fragment (SAN-18), 1 juvenile left humerus
diaphysis (SAN-38), 1 centrotarsal (SAN-37), 3 lower molars, either M1
or M2 (SAN-39, SAN-40 and SAN-41), and several fragments of
vertebras and ribs of difficult attribution to either species.
SAN-39
SAN-40
SAN-41
• This metacarpal is slender and quite small, similar to Bronze Age sheep
from the Iberian Peninsula. Using the factor proposed by Teichert (1975)
to calculate the height at the withers from the metacarpal bone, the height
of this specimen would be 54.1 cm. There is a significant decrease in the
height of sheep between the Chalcolithic and the Bronze Age.
Furthermore, the height of this sheep is within the range of Iron Age
sheep from the area.
11. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
Felines
Panthera pardus Felis catus
• The only remains identified as Panthera pardus are
a left upper carnasial SAN-2 and a tympanic
bulla SAN-26, thus the MNI is 1.
• After comparing it to other Upper Pleistocene
leopards from South Europe, we can conclude
that this premolar could belong to a female. The
macrowear shows that this animal was about 5-6
years old
• SAN-19 is a partial skull of a small felid. It is
fragmented and the rostrum is not preserved.
Because of the partial preservation, the only
measures that could be compared were the
greatest breadth of the occipital condyles and
the greatest breadth of the foramen magnum.
• Comparing SAN-19 to wild cat data, the Cova
dos Santos cat is much smaller than fossil or
modern wild cats from Europe, so most
probably it was a domestic cat, of unknown
antiquity
12. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
Bears
• The two species of bears present in the Iberian Peninsula during the Upper Pleistocene
and Holocene, Ursus arctos and Ursus spelaeus, have been found in this cave.
• We identified a total NMI of 4: one small female cave bear, one large male brown bear, and
a perinatal and a cub. The remains of young animals are common in cave bear dens, but we
cannot identify which species the remains from this cave belong to by their appearance
alone.
Ursus spelaeus Ursus arctos
13. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
ZooMS:
Zooarchaeology
by Mass Spectrometry
14. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
• Collagen is a highly conserved protein in the
different vertebrates groups, but with small
differences in the DNA sequence.
• These differences in DNA are translated into
small differences in the collagen amino acid
sequence.
• In this technique, the enzyme trypsin cuts the
protein in specific sites, forming smaller
peptides.
• The resulting peptides are identified by their
mass/charge in the MALDI-TOF mass
spectrometer.
• In this way, these peptides can be used as
molecular markers to identify taxa.
15. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
• 20 samples were selected for ZooMS.
• For this techique we used the collagen extrated for stable isotopes. Not all the different markers
are always obtained, which leads to the inability to accurately identify the genus or species.
Ursus sp.
25%
Rhinocerotidae
10%
Bos/Bison
20%
Equus sp.
5%
Sus scrofa
5%
Gallus sp.
5%
Capra sp.
5%
Ovis/Rupicapra
5%
Ovis/Rupicapra or
Cervus elaphus
5%
Bos/Bison, Cervus or
Ovis/Rupicapra
5%
Bos/Bison or
Ovis/Rupicapra
5%
Unidentified
5%
ZooMS results
16. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
SAN-09 SAN-15 SAN-34
SAN-36
SAN-45
• SAN-09, SAN-15, SAN-
34, SAN-36 and SAN-45
have been identified as
Ursus sp.
SAN-09
Genus Ursus
17. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
SAN-03 SAN-49
• SAN-03 and SAN-49 show markers compatible with
the family Rhinocerotidae. With the available
published markers it is not possible to distinguish the
genus.
• Two species inhabited the Iberian Peninsula during
the Upper Pleistocene: the woolly rhino (Coelodonta
antiquitatis) and the narrow-nosed rhino (Stephanorhinus
hemitoechus). The Iberian Peninsula could have acted
during MIS 3 as a glacial refuge for S. hemitoechus,
because it was not noticed in mainland Europe at this
time.
1199 1622 2869
2999
SAN-03
Family Rhinocerotidae
18. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
SAN-04
SAN-07
• Although the ZooMs
analysis does not distinguish
between Bos and Bison, the
morphological identification
of SAN-04 is Bos taurus, and
it is not excluded that the
rest of the bones identified
as Bos/Bison are also Bos
taurus.
SAN-27
SAN-46
Genera Bos or Bison
1209 1580
1648 3018
3033
SAN-07
19. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
SAN-14
SAN-14
1427
1550
2999
Genus Equus
Sus scrofa
SAN-16
SAN-16
1550
3017,3033
20. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
SAN-06 Capra sp.
SAN-10
Morphologically identified as Ovis aries
Bos/Bison or Cervus or Ovis/Rupicapra SAN-11 Bos/Bison or Ovis/Rupicapra
Ovicaprines
SAN-12 Ovis/Rupicapra
21. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
SAN-08
Ovis/Rupicapra or Cervus elaphus
Identified as Cervus elaphus because its larger size.
SAN-16
Unidentified
In this case, the peptides
obtained were not
enough to identify this
bone when compared
with existing databases.
23. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
• From the faunal assemblage, 4 samples were selected for 14C AMS dating: rhinoceros, leopard, red deer and
equid.
• The cave bear jaw SAN-1 falls
into the range of dates for the
youngest cave bears from Cova
Eirós, an important cave bear
den (located 120 km away).
• Although at a period of
marked climatic fluctuations,
the dating of the rhinoceros
coincides with a not
particularly cold phase.
Radiocarbon datings from Galicia (Cova dos Santos in bold dots):
• The leopard and the deer are the only fossils recovered up to now from the MIS 2 of Galicia. The leopard
dates from the beginning of the LGM, and the deer, in turn, dates back to the beginning of the temperate
pulse after the Heinrich 1 stadial.
• Finally, the 14C dating of the horse indicates a very recent age: 170 ± 30 yr BP which, once calibrated, yields
an age ranging from the eighteenth century to the present day.
25. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
• A total of 23 stable isotope analyses were performed.
• The main factor determining the isotopic signature in an
animal’s collagen is its diet type, but also various climatic
factors (temperature, humidity and insolation) produce
variations in the isotopic values of soils and vegetation,
which are transmitted along the food web. Temperature
correlates directly with δ15N values, as it is related to
microbial activity in soils, which increases or decreases in
parallel to ambient temperature. In turn, δ13C values
correlate directly with aridity, as this decreases the selective
uptake of 12CO2 by C3 plants during photosynthesis. The
dates obtained place the fauna studied from the end of the
Upper Pleistocene to recent times, and in this time interval
the climatic fluctuations were extreme.
• Pleistocene fauna: in our results we observed higher δ15N
values in the leopard than in the herbivores (cave bear, deer,
rhino), as expected for a carnivore. The δ13C values
obtained also allow us to identify C3 plants as the basis of
the food chain, which is common in temperate regions of
Europe.
• Domestic animals: the anthropic management of livestock and pastures may also interfere, mainly due to the
use of organic fertilizers which increase δ15N values as can be seen in the presumably domestic fauna
(chicken, pig, at least one of the cows).
26. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
Taphonomic interpretation of the faunal assemblage
• No features in the topography of the cave suggest the existence of an
ancient chasm into which the animals could have fallen. On the contrary, the
integration of the digital model of the cave in DEM terrain models shows
how there could potentially be an old entrance near the steep limit of some
nearby fields. The dimensions of the front of the filling point to the
possibility that there could have been a large entrance, which would have
facilitated both the fall of remains and the access of large vertebrates.
• The fragmental nature of the skeletal remains recovered and the confluence
of different species and chronologies show that the primary deposits were
removed and transported, with only a small portion of remains now found
on the surface, ranging from the end-Pleistocene fauna to recent domestic
animals.
• The presence of at least one adult female bear, a young bear and a perinatal
bear, as well as bear claw bioglyphs, suggest that the cave served as a
hibernation and breeding cave at least during the occupation by the cave
bear, dated to ca. 30.000 years ago. Scratches of juvenile claws on the cave
walls are frequent features in Galician caves with cave bear dens. The
presence of brown bears is also justified by their habit of hibernating in
caves. In this case, it is a large male brown bear.
27. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
Taphonomic interpretation of the faunal assemblage
• The rhinoceros remains are the oldest of the dated bones. There is no simple explanation for their presence
in the cave, unless they were transported from outside the cave by scavengers or by natural slope processes
at a time when the cave entrance was wider.
• The goat bone sample (NMI = 2) is very small, suggesting that it was a domestic animal and therefore
more recent than the leopard. The same is true for the cow and sheep, which may date from recent
prehistoric times to just a few centuries ago. Domestic fauna is frequent in the caves of NW Iberia, which
is related to accidental falls of animals in extensive rearing or to the use of the cave as a deposit for dead
animals. This group of domestic animals includes the equine, which is practically from the present day, the
chicken and possibly the pig and the cat.
• Leopard feeding remains are characterised by the
presence of poorly modified bones, with few tooth
marks and an absence of digested bones. In
European Pleistocene leopard dens, the most
common prey is the ibex and the chamois. In Cova
dos Santos, no digested bones were observed.
Bones with gnawing marks are scarce and the marks
are small and affecting basically the remains of
bears and ovicaprines.
28. Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)
Conclusions
• During the topography work in the Cova dos Santos, a superficial survey was carried out in
which a modest collection of bone remains was recovered, apparently of little value, given
their poor preservation and difficulty of identification in most cases.
• The bones are found in secondary deposits in several places in the cave, which must have had
a wider entrance in the past, as can be deduced from its topographical features.
• From 50 remains, mostly fragmentary, and combining classical and molecular palaeontological
techniques, it has been possible to identify animals of Pleistocene age such as the cave bear,
leopard, rhinoceros and deer. Also domestic animals such as cows, sheep, goat, chicken and
horse, and other animals of uncertain age such as the brown bear and a suid.
• The cave probably served as a cave bear den, a den for some other carnivore (perhaps
leopard) and as an accumulation of domestic mammals.
29. Thank you for your attention!
Molecular paleontology applied to sites with fragmented remains:
the case of Cova dos Santos (NW Spain)